Characterization of sucrose nonfermenting-1-related protein kinase 2 (SnRK2) gene family in Haynaldia villosa demonstrated SnRK2.9-V enhances drought and salt stress tolerance of common wheat.

BACKGROUND: The sucrose nonfermenting-1-related protein kinase 2 (SnRK2) plays a crucial role in responses to diverse biotic/abiotic stresses. Currently, there are reports on these genes in Haynaldia villosa, a diploid wild relative of wheat. RESULTS: To understand the evolution of SnRK2-V family genes and their roles in various stress conditions, we performed genome-wide identification of the SnRK2-V gene family in H. villosa. Ten SnRK2-V genes were identified and characterized for their structures, functions and spatial expressions. Analysis of gene exon/intron structure further revealed the presence of evolutionary paths and replication events of SnRK2-V gene family in the H. villosa. In addition, the features of gene structure, the chromosomal location, subcellular localization of the gene family were investigated and the phylogenetic relationship were determined using computational approaches. Analysis of cis-regulatory elements of SnRK2-V gene members revealed their close correlation with different phytohormone signals. The expression profiling revealed that ten SnRK2-V genes expressed at least one tissue (leave, stem, root, or grain), or in response to at least one of the biotic (stripe rust or powdery mildew) or abiotic (drought or salt) stresses. Moreover, SnRK2.9-V was up-regulated in H. villosa under the drought and salt stress and overexpressing of SnRK2.9-V in wheat enhanced drought and salt tolerances via enhancing the genes expression of antioxidant enzymes, revealing a potential value of SnRK2.9-V in wheat improvement for salt tolerance. CONCLUSION: Our present study provides a basic genome-wide overview of SnRK2-V genes in H. villosa and demonstrates the potential use of SnRK2.9-V in enhancing the drought and salt tolerances in common wheat.

Chromosome painting reveals inter-chromosomal rearrangements and evolution of subgenome D of wheat.

Aegilops species represent the most important gene pool for breeding bread wheat (Triticum aestivum). Thus, understanding the genome evolution, including chromosomal structural rearrangements and syntenic relationships among Aegilops species or between Aegilops and wheat, is important for both basic genome research and practical breeding applications. In the present study, we attempted to develop subgenome D-specific fluorescence in situ hybridization (FISH) probes by selecting D-specific oligonucleotides based on the reference genome of Chinese Spring. The oligo-based chromosome painting probes consisted of approximately 26 000 oligos per chromosome and their specificity was confirmed in both diploid and polyploid species containing the D subgenome. Two previously reported translocations involving two D chromosomes have been confirmed in wheat varieties and their derived lines. We demonstrate that the oligo painting probes can be used not only to identify the translocations involving D subgenome chromosomes, but also to determine the precise positions of chromosomal breakpoints. Chromosome painting of 56 accessions of Ae. tauschii from different origins led us to identify two novel translocations: a reciprocal 3D-7D translocation in two accessions and a complex 4D-5D-7D translocation in one accession. Painting probes were also used to analyze chromosomes from more diverse Aegilops species. These probes produced FISH signals in four different genomes. Chromosome rearrangements were identified in Aegilops umbellulata, Aegilops markgrafii, and Aegilops uniaristata, thus providing syntenic information that will be valuable for the application of these wild species in wheat breeding.

Recognition of glycoside hydrolase 12 proteins by the immune receptor RXEG1 confers Fusarium head blight resistance in wheat.

Fusarium head blight (FHB), caused by Fusarium graminearum, is a devastating disease in wheat (Triticum aestivum) that results in substantial yield losses and mycotoxin contamination. Reliable genetic resources for FHB resistance in wheat are lacking. In this study, we characterized glycoside hydrolase 12 (GH12) family proteins secreted by F. graminearum. We established that two GH12 proteins, Fg05851 and Fg11037, have functionally redundant roles in F. graminearum colonization of wheat. Furthermore, we determined that the GH12 proteins Fg05851 and Fg11037 are recognized by the leucine-rich-repeat receptor-like protein RXEG1 in the dicot Nicotiana benthamiana. Heterologous expression of RXEG1 conferred wheat responsiveness to Fg05851 and Fg11037, enhanced wheat resistance to F. graminearum and reduced levels of the mycotoxin deoxynivalenol in wheat grains in an Fg05851/Fg11037-dependent manner. In the RXEG1 transgenic lines, genes related to pattern-triggered plant immunity, salicylic acid, jasmonic acid, and anti-oxidative homeostasis signalling pathways were upregulated during F. graminearum infection. However, the expression of these genes was not significantly changed during infection by the deletion mutant ΔFg05851/Fg11037, suggesting that the recognition of Fg05851/Fg11037 by RXEG1 triggered plant resistance against FHB. Moreover, introducing RXEG1 into three other different wheat cultivars via crossing also conferred resistance to F. graminearum. Expression of RXEG1 did not have obvious deleterious effects on plant growth and development in wheat. Our study reveals that N. benthamiana RXEG1 remains effective when transferred into wheat, a monocot, which in turn suggests that engineering wheat with interfamily plant immune receptor transgenes is a viable strategy for increasing resistance to FHB.

Isoprenylation modification is required for HIPP1-mediated powdery mildew resistance in wheat.

Powdery mildew (Pm), caused by Blumeria graminis f.sp. tritici (Bgt), is one of the most important wheat diseases. Heavy-metal-associated isoprenylated plant protein (HIPP1) has been proved playing important roles in response to biotic and a biotic stress. In present study, we proved HIPP1-V from Haynalidia villosa is a positive regulator in Pm resistance. HIPP1-V was rapidly induced by Bgt. Transiently or stably heterologous overexpressing HIPP1-V in wheat suppressed the haustorium formation and enhanced Pm resistance. HIPP1-V isoprenylation was critical for plasma membrane (PM) localization, interaction with E3-ligase CMPG1-V and function in Pm resistance. Bgt infection recruited the isoprenylated HIPP1-V and CMPG1s on PM; blocking the HIPP1 isoprenylation reduced such recruitment and compromised the resistance of OE-CMPG1-V and OE-HIPP1-V. Overexpressing HIPP1-VC148G could not enhance Pm resistance. These indicated the Pm resistance was dependent on HIPP1-V's isoprenylation. DGEs related to the ROS and SA pathways were remarkably enriched in OE-HIPP1-V, revealing their involvement in Pm resistance. Our results provide evidence on the important role of protein isoprenylation in plant defense.

Fine mapping of two recessive powdery mildew resistance genes from Aegilops tauschii accession CIae8.

KEY MESSAGE: Two recessive powdery mildew resistance loci pmAeCIae8_2DS and pmAeCIae8_7DS from Aegilops tauschii were mapped and two synthesized hexaploid wheat lines were developed by distant hybridization. Wheat powdery mildew (Pm), one of the worldwide destructive fungal diseases, causes significant yield loss up to 30%. The identification of new Pm resistance genes will enrich the genetic diversity of wheat breeding for Pm resistance. Aegilops tauschii is the ancestor donor of sub-genome D of hexaploid wheat. It provides beneficial genes that can be easily transferred into wheat by producing synthetic hexaploid wheat followed by genetic recombination. We assessed the Pm resistance level of 35 Ae. tauschii accessions from different origins. Accession CIae8 exhibited high Pm resistance. Inheritance analysis and gene mapping were performed using F2 and F2:3 populations derived from the cross between CIae8 and a Pm susceptible accession PI574467. The Pm resistance of CIae8 was controlled by two independent recessive genes. Bulked segregate analysis using a 55 K SNP array revealed the SNPs were mainly enriched into genome regions, i.e. 2DS (13.5-20 Mb) and 7DS (4.0-15.5 Mb). The Pm resistance loci were named as pmAeCIae8_2DS and pmAeCIae8_7DS, respectively. By recombinant screening, we narrowed the pmAeCIae8_2DS into a 370-kb interval flanked by markers CINAU-AE7800 (14.89 Mb) and CINAU-AE20 (15.26 Mb), and narrowed the pmAeCIae8_7DS into a 260-kb interval flanked by markers CINAU-AE58 (4.72 Mb) and CINAU-AE25 (4.98 Mb). The molecular markers closely linked with the resistance loci were developed, and two synthesized hexaploid wheat (SHW) lines were produced. These laid the foundation for cloning of the two resistance loci and for transferring the resistance into common wheat.

Transferring a new Fusarium head blight resistance locus FhbRc1 from Roegneria ciliaris into wheat by developing alien translocation lines.

KEY MESSAGE: A new FHB resistance locus FhbRc1 was identified from the R. ciliaris chromosome 7Sc and transferred into common wheat by developing alien translocation lines. Fusarium head blight (FHB) caused by multiple Fusarium species is a globally destructive disease of common wheat. Exploring and utilization of resources with FHB resistance are the most effective and environmentally beneficial approach for the disease control. Roegneria ciliaris (Trin.) Nevski (2n = 4x = 28, ScScYcYc), a tetraploid wheat wild relative, possesses high resistance to FHB. In the previous study, a complete set of wheat-R. ciliaris disomic addition (DA) lines were evaluated for FHB resistance. DA7Sc had stable FHB resistance, which was confirmed to be derived from alien chromosome 7Sc. We tentatively designated the resistant locus as FhbRc1. For better utilization of the resistance in wheat breeding, we developed translocations by inducing chromosome structural aberrations using iron irradiation and the homologous pairing gene mutant ph1b. Totally, 26 plants having various 7Sc structural aberrations were identified. By marker analysis, a cytological map of 7Sc was constructed and 7Sc was dissected into 16 cytological bins. Seven alien chromosome aberration lines, which all had the bin 7Sc-1 on the long arm of 7Sc, showed enhanced FHB resistance. Thus, FhbRc1 was mapped to the distal region of 7ScL. A homozygous translocation line T4BS·4BL-7ScL (NAURC001) was developed. It showed improved FHB resistance, while had no obvious genetic linkage drag for the tested agronomic traits compared with the recurrent parent Alondra's. When transferring the FhbRc1 into three different wheat cultivars, the derived progenies having the translocated chromosome 4BS·4BL-7ScL all showed improved FHB resistance. This revealed the potential value of the translocation line in wheat breeding for FHB resistance.

Resistance of QYm.nau-2D to wheat yellow mosaic virus was derived from an alien introgression into common wheat.

KEY MESSAGE: The QYm.nau-2D locus conferring wheat yellow mosaic virus resistance is an exotic introgression and we developed 11 diagnostic markers tightly linked to QYm.nau-2D. Wheat yellow mosaic virus (WYMV) is a serious disease of winter wheat in China. Breeding resistant varieties is the most effective strategy for WYMV control. A WYMV resistant locus QYm.nau-2D on the chromosome arm 2DL has been repeatedly reported but the mapped region is large. In the present study, we screened recombinants using a biparental population and mapped QYm.nau-2D into an 18.8 Mb physical interval. By genome-wide association studies of 372 wheat varieties for WYMV resistance in four environments, we narrowed down QYm.nau-2D into a 16.4 Mb interval. Haplotype analysis indicated QYm.nau-2D were present as six different states due to recombination during hybridization breeding. QYm.nau-2D was finally mapped into a linkage block of 11.2 Mb. Chromosome painting using 2D specific probes and collinearity analysis among the published sequences corresponding to QYm.nau-2D region indicated the block was an exotic introgression. The Illumina-sequenced reads of four diploid Aegilops species were mapped to the sequence of Fielder, a variety having the introgression. The mapping reads were significantly increased at the putative introgression regions of Fielder. Ae. uniaristata (NN) had the highest mapping reads, suggesting that QYm.nau-2D was possibly an introgression from genome N. We investigated the agronomic performances of different haplotypes and observed no linkage drag of the alien introgression for the 15 tested traits. For marker-assisted selection of QYm.nau-2D, we developed 11 diagnostic markers tightly linked to the locus. This research provided a case study of an exotic introgression, which has been utilized in wheat improvement for WYMV resistance.

Dissection and cytological mapping of chromosome arm 4VS by the development of wheat-Haynaldia villosa structural aberration library.

KEY MESSAGE: A cytological map of Haynaldia villosa chromosome arm 4VS was constructed to facilitate the identification and utilization of beneficial genes on 4VS. Induction of wheat-alien chromosomal structure aberrations not only provides new germplasm for wheat improvement, but also allows assignment of favorable genes to define physical regions. Especially, the translocation or introgression lines carrying alien chromosomal fragments with different sizes are useful for breeding and alien gene mapping. Chromosome arm 4VS of Haynaldia villosa (L.) Schur (syn. Dasypyrum villosum (L.) P. Candargy) confers resistances to eyespot and wheat yellow mosaic virus (WYMV). In this research, we used both irradiation and the pairing homoeologous gene (Ph) mutant to induce chromosomal aberrations or translocations. By using the two approaches, a structural aberration library of chromosome arm 4VS was constructed. In this library, there are 57 homozygous structural aberrations, in which, 39 were induced by the Triticum aestivum cv. Chinese Spring (CS) ph1b mutant (CS ph1b) and 18 were induced by irradiation. The aberrations included four types, i.e., terminal translocation, interstitial translocation, deletion and complex structural aberration. The 4VS cytological map was constructed by amplification in the developed homozygous aberrations using 199 4VS-specific markers, which could be allocated into 39 bins on 4VS. These bins were further assigned to their corresponding physical regions of chromosome arm 4DS based on BLASTn search of the marker sequences against the reference sequence of Aegilops tauschii Cosson. The developed genetic stocks and cytological map provide genetic stocks for wheat breeding as well as alien gene tagging.

The Regulatory Network of CMPG1-V in Wheat-Blumeria graminis f. sp. tritici Interaction Revealed by Temporal Profiling Using RNA-Seq.

Wheat powdery mildew (Pm), caused by Blumeria graminis f. sp. tritici (Bgt), is a prevalent fungal disease. The diploid wheat relative Haynaldia villosa (H. villosa) showed broad-spectrum resistance (BSR) to Pm. A previous study reported an E3 ligase gene, CMPG1-V from H. villosa, showing BSR to Pm. To elucidate the regulatory network mediated by CMPG1-V, in this study, gene expression profiling of CMPG1-V transgenic plant (CMPG1-VOE) and its receptor Yangmai 158 was analyzed and compared after Bgt inoculation at four infection stages. GO and KEGG analysis revealed obvious reprogramming of SA and ABA signaling, starch/sucrose metabolism, and photosynthesis in CMPG1-VOE, compared with those in Yangmai 158. Transcripts of SA synthesis genes SARD1 and UGT, signaling factors TGA and PRs, and SnRKs in ABA signaling were specifically upregulated in CMPG1-VOE rather than Yangmai 158. Transcripts of LHCII in photosynthesis, GLUC and TPP in starch/sucrose metabolism were also induced distinctly in CMPG1-VOE. WGCNA analysis showed crucial regulatory candidates of CMPG1-V, involving serine/threonine-protein kinase in phosphorylation, glucosyltransferase in flavonoid biosynthesis, defense factor WRKYs, and peroxidase in oxidative stress. Our results facilitate the deciphering of the resistant regulatory network of CMPG1-V and the identification of key candidates which might be employed in breeding programs.

LecRK-V, an L-type lectin receptor kinase in Haynaldia villosa, plays positive role in resistance to wheat powdery mildew.

Plant sense potential microbial pathogen using pattern recognition receptors (PRRs) to recognize pathogen-associated molecular patterns (PAMPs). The Lectin receptor-like kinase genes (LecRKs) are involved in various cellular processes mediated by signal transduction pathways. In the present study, an L-type lectin receptor kinase gene LecRK-V was cloned from Haynaldia villosa, a diploid wheat relative which is highly resistant to powdery mildew. The expression of LecRK-V was rapidly up-regulated by Bgt inoculation and chitin treatment. Its transcript level was higher in the leaves than in roots, culms, spikes and callus. Single-cell transient overexpression of LecRK-V led to decreased haustorium index in wheat variety Yangmai158, which is powdery mildew susceptible. Stable transformation LecRK-V into Yangmai158 significantly enhanced the powdery mildew resistance at both seedling and adult stages. At seedling stage, the transgenic line was highly resistance to 18 of the tested 23 Bgt isolates, hypersensitive responses (HR) were observed for 22 Bgt isolates, and more ROS at the Bgt infection sites was accumulated. These indicated that LecRK-V confers broad-spectrum resistance to powdery mildew, and ROS and SA pathways contribute to the enhanced powdery mildew resistance in wheat.

E3 ubiquitin ligase gene CMPG1-V from Haynaldia villosa L. contributes to powdery mildew resistance in common wheat (Triticum aestivum L.).

Powdery mildew is one of the most devastating wheat fungal diseases. A diploid wheat relative, Haynaldia villosa L., is highly resistant to powdery mildew, and its genetic resource of resistances, such as the Pm21 locus, is now widely used in wheat breeding. Here we report the cloning of a resistance gene from H. villosa, designated CMPG1-V, that encodes a U-box E3 ubiquitin ligase. Expression of the CMPG1-V gene was induced in the leaf and stem of H. villosa upon inoculation with Blumeria graminis f. sp. tritici (Bgt) fungus, and the presence of Pm21 is essential for its rapid induction of expression. CMPG1-V has conserved key residues for E3 ligase, and possesses E3 ligase activity in vitro and in vivo. CMPG1-V is localized in the nucleus, endoplasmic reticulum, plasma membrane and partially in trans-Golgi network/early endosome vesicles. Transgenic wheat over-expressing CMPG1-V showed improved broad-spectrum powdery mildew resistance at seedling and adult stages, associated with an increase in expression of salicylic acid-responsive genes, H2 O2 accumulation, and cell-wall protein cross-linking at the Bgt infection sites, and the expression of CMPG1-V in H. villosa was increased when treated with salicylic acid, abscisic acid and H2 O2 . These results indicate the involvement of E3 ligase in defense responses to Bgt fungus in wheat, particularly in broad-spectrum disease resistance, and suggest association of reactive oxidative species and the phytohormone pathway with CMPG1-V-mediated powdery mildew resistance.

Dosage effect genes modulate grain development in synthesized Triticum durum-Haynaldia villosa allohexaploid.

Polyploidization in plants often leads to increased cell size and grain size, which may be affected by the increased genome dosage and transcription abundance. The synthesized Triticum durum (AABB)-Haynaldia villosa (VV) amphiploid (AABBVV) has significantly increased grain size, especially grain length, than the tetraploid and diploid parents. To investigate how polyploidization affects grain development at the transcriptional level, we perform transcriptome analysis using the immature seeds of T. durum, H. villosa, and the amphiploid. The dosage effect genes are contributed more by differentially expressed genes from genome V of H. villosa. The dosage effect genes overrepresent grain development-related genes. Interestingly, the vernalization gene TaVRN1 is among the positive dosage effect genes in the T. durum‒H. villosa and T. turgidum‒Ae. tauschii amphiploids. The expression levels of TaVRN1 homologs are positively correlated with the grain size and weight. The TaVRN1-B1 or TaVRN1-D1 mutation shows delayed florescence, decreased cell size, grain size, and grain yield. These data indicate that dosage effect genes could be one of the important explanations for increased grain size by regulating grain development. The identification and functional validation of dosage effect genes may facilitate the finding of valuable genes for improving wheat yield.

A chromosome-scale genome assembly of Dasypyrum villosum provides insights into its application as a broad-spectrum disease resistance resource for wheat improvement.

Dasypyrum villosum is one of the most valuable gene resources in wheat improvement, especially for disease resistance. The mining of favorable genes from D. villosum is frustrated by the lack of a whole genome sequence. In this study, we generated a doubled-haploid line, 91C43DH, using microspore culture and obtained a 4.05-GB high-quality, chromosome-scale genome assembly for D. villosum. The assembly contains39 727 high-confidence genes, and 85.31% of the sequences are repetitive. Two reciprocal translocation events were detected, and 7VS-4VL is a unique translocation in D. villosum. The prolamin seed storage protein-coding genes were found to be duplicated; in particular, the genes encoding low-molecular-weight glutenin at the Glu-V3 locus were significantly expanded. RNA sequencing (RNA-seq) analysis indicated that, after Blumeria graminearum f.sp tritici (Bgt) inoculation, there were more upregulated genes involved in the pattern-triggered immunity and effector-triggered immunity defense pathways in D. villosum than in Triticum urartu. MNase hypersensitive sequencing (MH-seq) identified two Bgt-inducible MH sites (MHSs), one in the promoter and one in the 3' terminal region of the powdery mildew resistance (Pm) gene NLR1-V. Each site had two subpeaks and they were termed MHS1 (MHS1.1/1.2) and MHS2 (MHS2.1/2.2). Bgt-inducible MHS2.2 was uniquely present in D. villosum, and MHS1.1 was more inducible in D. villosum than in wheat, suggesting that MHSs may be critical for regulation of NLR1-V expression and plant defense. In summary, this study provides a valuable genome resource for functional genomics studies and wheat-D. villosum introgression breeding. The identified regulatory mechanisms may also be exploited to develop new strategies for enhancing Pm resistance by optimizing gene expression in wheat.

Pleiotropic Effect of the compactum Gene and Its Combined Effects with Other Loci for Spike and Grain-Related Traits in Wheat.

Club wheat (Triticum aestivum ssp. compactum) with a distinctly compact spike morphology was conditioned by the dominant compactum (C) locus on chromosome 2D and resulted in a redistribution of spike yield components. The disclosure of the genetic basis of club wheat was a prerequisite for the development of widely adapted, agronomically competitive club wheat cultivars. In this study, we used a recombinant inbred line population derived from a cross between club wheat Hiller and modern cultivar Yangmai 158 to construct a genetic linkage map and identify quantitative trait loci associated with 15 morphological traits. The club allele acted in a semi-dominant manner and the C gene was mapped to 370.12-406.29 Mb physical region on the long arm of 2D. Apart from compact spikes, C exhibited a pleiotropic effect on ten other agronomic traits, including plant height, three spike-related traits and six grain-related traits. The compact spike phenotype was correlated with decreased grain size and weight, but with an increase in floret fertility and grain number. These pleiotropic effects make club wheat have compatible spike weight with a normal spike from common wheat. The genetic effects of various gene combinations of C with four yield-related genes, including Ppd-D1, Vrn-D3, Rht-B1b and Rht8, were evaluated. C had no epistatic interaction with any of these genes, indicating that their combinations would have an additive effect on other agronomically important traits. Our research provided a theoretical foundation for the potentially effective deployment of C gene into modern breeding varieties in combination with other favorable alleles.

The Exocyst Complex Subunit EXO70E1-V From Haynaldia villosa Interacts With Wheat Powdery Mildew Resistance Gene CMPG1-V.

EXO70 belongs to the exocyst complex subunit that plays a critical role in regulating plant cell polarity establishment and defense response. A previous study proved that the E3 ligase CMPG1-V from Haynaldia villosa, a diploid wheat relative, positively regulates the resistance to wheat powdery mildew (Pm), caused by fungus Blumeria graminis f.sp tritici (Bgt). In this study, a member of EXO70 superfamily named EXO70E1-V was isolated from H. villosa, and EXO70E1-V interacted with CMPG1-V were shown by yeast two-hybrid (Y2H), pull-down assay, bimolecular fluorescence complementation (BiFC) assay, and luciferase complementation imaging (LCI) assay. It is localized in various subcellular organs, i.e., plasma membrane (PM) and endoplasmic reticulum. Co-expression of EXO70E1-V and CMPG1-V showed dot-like structure fluorescence signals that were mainly in PM and nucleus. Expression of EXO70E1-V was relatively higher in leaf and was significantly induced by Bgt infection and exogenous application of hormones such as salicylic acid. Transient or stable overexpression of EXO70E1-V could not enhance/decrease the Pm resistance level, suggesting overexpression of EXO70E1-V alone has no impact on Pm resistance in wheat.

Discovery of broad-spectrum fungicides that block septin-dependent infection processes of pathogenic fungi.

Many pathogenic fungi depend on the development of specialized infection structures called appressoria to invade their hosts and cause disease. Impairing the function of fungal infection structures therefore provides a potential means by which diseases could be prevented. In spite of this extraordinary potential, however, relatively few anti-penetrant drugs have been developed to control fungal diseases, of either plants or animals. In the present study, we report the identification of compounds that act specifically to prevent fungal infection. We found that the organization of septin GTPases, which are essential for appressorium-mediated infection in the rice blast fungus Magnaporthe oryzae, requires very-long-chain fatty acids (VLCFAs), which act as mediators of septin organization at membrane interfaces. VLCFAs promote septin recruitment to curved plasma membranes and depletion of VLCFAs prevents septin assembly and host penetration by M. oryzae. We observed that VLCFA biosynthesis inhibitors not only prevent rice blast disease, but also show effective, broad-spectrum fungicidal activity against a wide range of fungal pathogens of maize, wheat and locusts, without affecting their respective hosts. Our findings reveal a mechanism underlying septin-mediated infection structure formation in fungi and provide a class of fungicides to control diverse diseases of plants and animals.

A disulphide isomerase gene (PDI-V) from Haynaldia villosa contributes to powdery mildew resistance in common wheat.

In this study, we report the contribution of a PDI-like gene from wheat wild relative Haynaldia villosa in combating powdery mildew. PDI-V protein contains two conserved thioredoxin (TRX) active domains (a and a') and an inactive domain (b). PDI-V interacted with E3 ligase CMPG1-V protein, which is a positive regulator of powdery mildew response. PDI-V was mono-ubiquitinated by CMPG1-V without degradation being detected. PDI-V was located on H. villosa chromosome 5V and encoded for a protein located in the endoplasmic reticulum. Bgt infection in leaves of H. villosa induced PDI-V expression. Virus induced gene silencing of PDIs in a T. durum-H. villosa amphiploid compromised the resistance. Single cell transient over-expression of PDI-V or a truncated version containing the active TXR domain a decreased the haustorial index in moderately susceptible wheat cultivar Yangmai 158. Stable transgenic lines over-expressing PDI-V in Yangmai 158 displayed improved powdery mildew resistance at both the seedling and adult stages. By contrast over-expression of point-mutated PDI-V(C57A) did not increase the level of resistance in Yangmai 158. The above results indicate a pivotal role of PDI-V in powdery mildew resistance and showed that conserved TRX domain a is critical for its function.